Guanylin and uroguanylin are peptides that are expressed in intestinal epithelia and were identified because of their ability to bind to the transmembrane receptor, guanylate cyclase C (GC-C). Ligand binding to GC-C, also expressed on intestinal epithelial cells (IEC), elevates intracellular cGMP levels which in turn elicits transmembrane ion movement via the cystic fibrosis transmembrane conductance regulator and the Na+/H+ Exchanger 3. This signaling pathway has direct clinical relevance in that the acute diarrheal illness caused by enterotoxigenic strains of E. coli is mediated by the heat stable toxin, a super-agonist of GC-C. However, receptor guanylate cyclases that are highly homologous to GC-C are important in regulating pro-inflammatory gene expression, barrier function, and inflammation in extraintestinal organs. While a significant amount of published work has centered on GC-C regulation of fluid secretion, almost nothing has been reported regarding the putative role for GC-C in intestinal inflammatory disease. Moreover, recent findings indicate that GC-C ligands are reduced in patients with Inflammatory Bowel Disease. Therefore, a better understanding of this receptor-ligand system will likely shed light not only on the pathophysiology of toxigenic diarrhea, but will also serve to close the current gap in our understanding of GC-C and its ligands in the pathogenesis of colitis. Consistent with this goal, our preliminary data indicates that signaling through GC-C has an important role in suppressing pro-inflammatory gene expression in intestinal mucosa, and is required for intestinal barrier function. Further, we demonstrate an intriguing differential sensitivity to innate immune cell-driven intestinal injury versus T cell-mediated colitis. Therefore, the overarching hypothesis of this application is that GC-C, via ligand-induced cGMP generation, is essential for mucosal immune homeostasis. We will address this in the following aims. Specific Aim 1 will investigate GC-C-mediated suppression of pro-inflammatory gene expression using in vitro biochemical analysis and existing, novel gene-targeted mice. Specific Aim 2 will evaluate the requirement for GC-C signaling in maintaining intestinal barrier function through the use of in vitro IEC monolayers and GC-C and ligand knockout animals. Specific Aim 3 will investigate the role of GC-C in susceptibility to intestinal inflammation using multiple in vivo models of colitis in GC-C null mice. The studies in this proposal will define the role of GC-C and its ligands in mucosal pro-inflammatory gene expression and barrier function, and will delineate the impact GC-C has on susceptibility to intestinal inflammation. Furthermore, these studies will lay the groundwork for translational research involving this receptor as a therapeutic target for the treatment of intestinal inflammatory disorders.